Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained ImmunogenicityReport as inadecuate

Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity - Download this document for free, or read online. Document in PDF available to download.

World Allergy Organization Journal

, Volume 4, Issue 7, pp 113–120

First Online: 14 July 2011


BackgroundRecombinant allergens are under investigation for replacing allergen extracts in immunotherapy. Site-directed mutagenesis has been suggested as a strategy to develop hypoallergenic molecules that will reduce the risk of side effects. For decades, chemically modified allergen extracts have been used for the same reason.

AimTo evaluate whether glutaraldehyde modification is a good strategy to produce hypoallergenic recombinant allergens with retained immunogenicity.

MethodsFel d 1 was cloned as a single construct linking both chains of the molecule and expressed in Escherichia coli and Pichia pastoris. After physicochemical purification, recombinant r Fel d 1 was chemically modified using glutaraldehyde. The effect of modification on immune reactivity was evaluated using radioallergosorbent test, CAP inhibition, competitive radioimmunoassay, enzyme-linked immunosorbent assay, basophil histamine release, and T-cell proliferation assays. Both natural and recombinant unmodified Fel d 1 were used as controls.

ResultsrFel d 1 demonstrated similar IgE binding and biologic activity as its natural counterpart. Upon modification, IgE-binding potency decreased >1000-fold, translating into a >10-fold reduction in biologic activity assessed by basophil histamine release. In contrast, the modified recombinant did not show a decreased but even a moderately increased capacity 1.5-fold to stimulate proliferation of T cells P < 0.01. Finally, it induced specific IgG antibodies in rabbits that recognized the unmodified allergen.

ConclusionsChemical modification is a practical and highly effective approach for achieving hypoallergenicity of recombinant allergens with retained immunogenicity.

Keywordsallergoid Felis domesticus hypoallergen immunotherapy rFel d 1  Download fulltext PDF

Author: Serge A Versteeg - Ingrid Bulder - Martin Himly - Toni M van Capel - R van den Hourt - Stef J Koppelman - Esther C de J


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